Genetic Engineering of Mammalian Sensory Circuits
哺乳动物感觉回路的基因工程
基本信息
- 批准号:RGPIN-2015-04876
- 负责人:
- 金额:$ 2.4万
- 依托单位:
- 依托单位国家:加拿大
- 项目类别:Discovery Grants Program - Individual
- 财政年份:2016
- 资助国家:加拿大
- 起止时间:2016-01-01 至 2017-12-31
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
Optogenetics has emerged as a powerful tool to control neuronal activity with high spatio-temporal precision. We were the first to develop optogenetic strategies to activate selectively genetically-defined peripheral somatosensory neurons involved in nociception and pain in mice (Daou et al., 2013). In this 5-year research program, we propose to design and validate conditional transgenic models for the functional interrogation of neuronal subsets with unclear roles in somatosensory circuits.Our main scientific objectives are:
1) To interrogate genetically-identified neurons in somatosensory circuits in vivo using optogenetics. Primary sensory neurons are located in the dorsal root, trigeminal, and nodose ganglia while their peripheral terminals and fibers innervate the skin and peripheral tissues. Their activation is directly linked to somatosensory modalities (touch, temperature, itch, nociception) and to pain perception. We previously used the "proof-of-concept" Nav1.8-Cre driver with broad neuronal expression but we propose now to interrogate more specific subsets of genetically-defined neurons: thermal peptidergic nociceptors with TRPV1-Cre, tactile non-peptidergic nociceptors with MrgD-Cre, itch sensors with MrgA3-Cre and mechanosensors with TH-Cre. Once their optical activation with ChR2 or inhibition with Arch is demonstrated in vitro and in vivo behaviorally in normal conditions (tactile response, thermal sensitivity, conditioned place aversion), we will test their contribution to chronic pain following inflammation or neuropathic injury using behavioral analysis and quantitative sensory measurements.
2) To modulate selectively the two main types of nociceptors in vivo using chemogenetics. Peripheral sensitization of nociceptors plays a major role in triggering central mechanisms of chronic pain. We have shown that TRPC3-linked Gq-coupled pathways contribute to peripheral sensitization in nociceptors (Alkhani et al., 2014). Using AAV viral expression of two chemogenetic actuators, the DREADDs M3D (Gq-coupled) and M4D (Gi-coupled) activated by the compound CNO, we propose to investigate the functional consequences of CNO-induced modulation of TRPV1+ and MrgD+ nociceptor firing in transgenic mice in vitro and in vivo. We make the hypothesis that activation of M3D Gq-coupled pathways will promote sensitization and inflammation, while M4D Gi-coupled pathways will protect DRG/TG neurons from sensitization and hyperexcitability, recruiting cellular mechanisms analogous to morphine analgesia.
These novel transgenic mice engineered with artificial peripheral transducers/receptors will allow us to dissect in vitro the signaling diversity of sensory neurons as well as to investigate in vivo the contribution of specific neuronal populations to several modalities of somatosensation relevant to pain, in normal and sensitized states.
光遗传学已经成为以高时空精度控制神经元活动的有力工具。我们是第一个开发光遗传策略来激活涉及小鼠伤害性感觉和疼痛的选择性遗传定义的外周体感神经元的策略(Daou等人,2013年)。在这个为期5年的研究计划中,我们建议设计和验证条件转基因模型,用于对躯体感觉回路中角色不明的神经元亚群进行功能询问。我们的主要科学目标是:
1)利用光遗传学在活体体感回路中询问遗传识别的神经元。初级感觉神经元位于背根、三叉神经节和结状神经节,而它们的外周终末和纤维分布于皮肤和周围组织。它们的激活直接与躯体感觉方式(触摸、温度、瘙痒、伤害感)和痛觉有关。我们之前使用了具有广泛神经元表达的“概念验证”Nav1.8-CRE驱动程序,但现在我们建议询问更特定的遗传定义神经元子集:具有TRPV1-CRE的热肽能伤害性感受器,具有MRgD-CRE的触觉非肽能伤害性感受器,具有MRgA3-CRE的瘙痒传感器和具有TH-CRE的机械传感器。一旦它们被ChR2激活或被Arch抑制,在体外和体内正常情况下的行为(触觉反应、热敏感性、条件性位置厌恶),我们将使用行为分析和定量感觉测量来测试它们对炎症或神经病理性损伤后慢性疼痛的贡献。
2)利用化学遗传学在体内选择性地调节两种主要类型的伤害性感受器。伤害性感受器的外周敏化在触发慢性疼痛的中枢机制中起主要作用。我们已经证明,TRPC3连接的GQ耦合通路有助于伤害性感受器的外周敏化(Alkhani等人,2014年)。利用AAV病毒表达的两种化学致癌因子,DREADDS M3D(GQ偶联)和M4D(GI偶联)被化合物CNO激活,我们建议在体外和体内研究CNO诱导的TRPV1+和MRgD+伤害性感受器激活的功能后果。我们假设M3DGQ偶联通路的激活将促进敏化和炎症,而M4DGI偶联通路将保护DRG/TG神经元免受敏化和过度兴奋,从而招募类似于吗啡镇痛的细胞机制。
这些利用人工外周转导/受体进行工程的新型转基因小鼠将使我们能够在体外解剖感觉神经元的信号多样性,并在体内研究特定神经元群体在正常和致敏状态下对几种与疼痛相关的躯体感觉方式的贡献。
项目成果
期刊论文数量(0)
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Séguéla, Philippe其他文献
Séguéla, Philippe的其他文献
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{{ truncateString('Séguéla, Philippe', 18)}}的其他基金
Genetic engineering of mammalian somatosensory circuits
哺乳动物体感回路的基因工程
- 批准号:
RGPIN-2020-04368 - 财政年份:2022
- 资助金额:
$ 2.4万 - 项目类别:
Discovery Grants Program - Individual
Genetic engineering of mammalian somatosensory circuits
哺乳动物体感回路的基因工程
- 批准号:
RGPIN-2020-04368 - 财政年份:2021
- 资助金额:
$ 2.4万 - 项目类别:
Discovery Grants Program - Individual
Genetic engineering of mammalian somatosensory circuits
哺乳动物体感回路的基因工程
- 批准号:
RGPIN-2020-04368 - 财政年份:2020
- 资助金额:
$ 2.4万 - 项目类别:
Discovery Grants Program - Individual
Genetic Engineering of Mammalian Sensory Circuits
哺乳动物感觉回路的基因工程
- 批准号:
RGPIN-2015-04876 - 财政年份:2017
- 资助金额:
$ 2.4万 - 项目类别:
Discovery Grants Program - Individual
Genetic Engineering of Mammalian Sensory Circuits
哺乳动物感觉回路的基因工程
- 批准号:
RGPIN-2015-04876 - 财政年份:2015
- 资助金额:
$ 2.4万 - 项目类别:
Discovery Grants Program - Individual
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